Metal-catalyzed oxidation of proteins is believed to play an important role in physiopathology and aging. Oxidative damage includes introduction of carbonyl functions to certain amino acid side chains. Carbonyls can be conveniently measured spectrophotometrically after their reaction with dinitrophenylhydrazine. Using this method, our laboratory has shown in the past an increase of the carbonyl content of whole intracellular proteins with increased age, both in animals and humans. However, the precise nature of the carbonyl-bearing oxidation products is not known. The present project is aimed at the chemical characterization of such products, the development of analytical techniques for their measurement, and the application of these to the analysis of tissues and individual proteins. From previous research, we know that arginine and proline are oxidized to glutamic and lysine to adipic semialdehydes. At the present time, an analytical method for glutamic semialdehyde has been fully developed, while one for adipic semialdehyde is at an advanced stage. Both compounds are measured in protein hydrolysates by selected ion monitoring gas chromatography/mass spectrometry after isotopic dilution. Samples are first reduced to the hydrolysis stable forms, hydroxyaminovaleric acid (HAVA) and hydroxyaminocaproic acid (HACA), respectively, which are then converted to their trifluoroacetyl-methyl ester volatile derivatives; synthesis and calibration of natural abundance and deuterated standards has been completed for HAVA, which has been measured in glutamine synthetase oxidized in vitro and in rat liver proteins. Preliminary results indicate that, in fact, HAVA accounts for a substantial proportion of carbonyl groups in artificially oxidized glutamine oxidase. The qualitative and quantitative information gained through these studies will be useful to better understand the nature of the oxidative modifications, the extent of damage to individual proteins, and its bearing to physiopathology and aging.